Design, synthesis and anticancer biological evaluation of novel 1,4-diaryl-1,2,3-triazole retinoid analogues of tamibarotene (AM80)

Article abstract of DOI:10.21577/0103-5053.20170119

We report herein the design and synthesis via click chemistry of twelve novel triazole retinoid analogues of tamibarotene (AM80) and the evaluation of their anticancer activities against six cancer cell lines: HL60, K562, 786, HT29, MCF7 and PC3. Among the synthesized compounds, two were more potent than tamibarotene against solid tumor cells, and one of them had similar potency to tamibarotene against HL60 cells. The bioisosteric exchange between the amide group and the 1,2,3-triazole core in the retinoid agent tamibarotene (AM80) reported in this work is a valid strategy for the generation of useful compounds against cancer.

Full text of DOI:10.21577/0103-5053.20170119

http://dx.doi.org/10.21577/0103-5053.20170119
J. Braz. Chem. Soc., Vol. 29, No. 1, 109-124, 2018.
Printed in Brazil - ©2018 Sociedade Brasileira de Química
Article
Design, Synthesis and Anticancer Biological Evaluation of Novel 1,4-Diaryl-
1,2,3-triazole Retinoid Analogues of Tamibarotene (AM80)
Mariana A. A. Aleixo,a Taís M. Garcia,a Diego B. Carvalho,a Luiz H. Viana,a
Marcos S. Amaral,a Najla M. Kassab,a Marilin C. Cunha,b Indiara C. Pereira,b
Palimécio G. Guerrero Jr.,c Renata T. Perdomo,b Maria F. C. Matosb and
Adriano C. M. Baroni*,a
aLaboratório de Síntese e Química Medicinal (LASQUIM) and
bLaboratório de Biologia molecular e Cultura de Células,
Faculdade de Ciências Farmacêuticas, Alimentos e Nutrição (FACFAN),
Universidade Federal do Mato Grosso do Sul (UFMS), 79070-900 Campo Grande-MS, Brazil
cDepartamento de Química e Biologia (DAQBi),
Universidade Tecnológica Federal do Paraná (UTFPR), 80230-901 Curitiba-PR, Brazil
We report herein the design and synthesis via click chemistry of twelve novel triazole retinoid
analogues of tamibarotene (AM80) and the evaluation of their anticancer activities against six
cancer cell lines: HL60, K562, 786, HT29, MCF7 and PC3. Among the synthesized compounds,
two were more potent than tamibarotene against solid tumor cells, and one of them had similar
potency to tamibarotene against HL60 cells. The bioisosteric exchange between the amide group
and the 1,2,3-triazole core in the retinoid agent tamibarotene (AM80) reported in this work is a
valid strategy for the generation of useful compounds against cancer.
Keywords: 1,2,3-triazole retinoids, tamibarotene, click chemistry, bioisosterism, anticancer
activities
Introduction
subtypes: α, β and γ.3-5 RARα receptor have important
role in hematopoiesis. Agonists of RARα receptor are
used against acute promyelocytic leukemia (APL), and
in the chemoprevention of estrogen receptor-positive
(ER-positive) breast cancer cell line. Studies have showed
that normal RARβ signaling is important factor in the
control of certain types of cancers. RARγ receptor, for
instance, is important for the skin functioning and their
agonists have been used in dermatology for the treatment
of acne, pysoriasis and photodamaged skin.6 RXR
receptors have important role in metabolic diseases, as
type II diabetes. Their agonists have also been used in
the chemoprevention of breast cancer.6
Among several retinoid anticancer compounds, here we
consider the pan-RAR agonists ATRA 1 (natural ligand)3-5
and TTNPB 2 (synthetic ligand),3-5 the pan-RXR agonist
bexarotene 3,3-9 the selective RARα receptor synthetic
agonists AM80 43-9 and AM580 5,3-9 and UVI2007 6, an
RARβ agonist10 (Figure 1).
In recent years, cancers have been responsible for
8.2 million human deaths worldwide.1,2 The rising demand
for effective and safer anticancer drugs has led several
research groups to develop new strategies to synthesize a
wide range of anticancer molecules and to evaluate their
biological anticancer activities.2
Retinoids are class of chemical compounds that are
derivatives of vitamin A with a large number of biological
processes.3 Retinoids were originally developed to treat
skin disorders, but these compounds have other potential
therapeutic uses, such as in type II diabetes, viral infection,
metabolic diseases, Alzheimer’s disease, and, primarily,
cancer, due to their effects on growth differentiation and
apoptosis.3-5
The biological effects of retinoids result from their
modulation of retinoic acid receptors (RARs) and
retinoic X receptors (RXRs), each having three target
The molecular structures of retinoids can be divided
*e-mail: adriano.baroni@ufms.br
into three parts: a hydrophobic region, a linker unit, and

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Design, Synthesis and Anticancer Biological Evaluation of Novel 1,4-Diaryl-1,2,3-triazole
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Figure 1. Classical retinoids reported in the literature.
Figure 2. Design of retinoids containing a 1,2,3-triazole core.
a polar terminus (Figure 2). Molecular modifications3-10
can provide information about the structure-activity
relationships (SARs) of these compounds.
tamibarotene 4 being less toxic than ATRA, side effects
still have limited its use in the long-term treatment against
APL cancer.13,14
The introduction of an amide group in the linker
unit during the synthesis of tamibarotene (AM80) 4 and
AM580 5 has yielded compounds with increased selectivity
for the RARα receptor, because the amide group does
hydrogen bond interactions with the Ser232 residue.3,7
In comparison, ATRA 1 and TTNPB 2 (Figure 1), which
contains a nonpolar vinyl linker, do not interact with
Ser 232; therefore, these compounds can be considered
pan-RAR agonists.3,7
Compounds 1 and 4 were identified as effective anti-
cancer agents for the treatment of acute promyelocytic
leukemia (APL) by inducing the differentiation or
inhibition of cell proliferation. As a consequence of
these powerful biological effects, all-trans retinoic acid
(ATRA) and tamibarotene 4 have been used clinically to
treat APL.11-14 However, tamibarotene (AM80) 4 is ten
times more potent than ATRA in inducing differentiation
of HL60 and NB-4 cells, and it has lower drug resistance
against APL cancer.13,14 Tamibarotene 4 has fewer side
effects on the dermal epithelium than ATRA because
it does not bind to the RARγ receptor. Although
Considering our research group’s interest in synthesizing
novel anti-cancer agents, we focused our studies on the
development of novel retinoid molecules based on the
tamibarotene structure.
The compounds were designed using classical
modification strategies, such as the use of bioisosterism
(Figure 2). The 1,2,3-triazole rings are effective amide
surrogates due to their strong dipole moments, their
similarities in terms of distance and planarity, and also, as the
amides, 1,2,3-triazole core can make hydrogen bonds.15 We
chose 1,2,3-triazole moiety because it have also been found in
anticancer substances,16 and it is more metabolic stable than
amines.15 Thus, we believe that novel 1,2,3-triazole retinoids
designed herein, also may be a selective RARα agonist as
tamibarotene 4, and this approach may contribute to obtain
new derivatives with improved anticancer activities.
The carboxylic acid unit was also modified to obtain
SAR information for 7-17 (Figure 2).17 The unknown
1,5-diaryl-1,2,3-triazolic retinoid derivative 18 was
designed with the goal of comparing its anticancer activities
with those of compound 7.

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Results and Discussion
1,1,4,4-tetramethyl-1,2,3,4-tetrahydronaphthalene 22
in 82% yield,6,20 which was then reacted with an
NBS/TsOH/CH3OH system for 48 h under reflux to produce
bromotetrahydronaphthalene 23 (Scheme 1).21
Subsequently, a Sonogashira cross-coupling reaction
between bromide 23 and 2-methyl-3-butyn-2-ol 24
using PdCl2(PPh3)2 and CuI as cocatalysts and Et3N
as a base produced the acetylenic alcohol 25 in 81%
yield.20,22
The retro-Favorskii reaction of 25 with KOH under
reflux in toluene generated the terminal acetylene 26
with 70% yield.20,22 Next, the aromatic azides 28a-c were
prepared by the reaction of aromatic amines 27a-c with
t-BuONO/TMSN3 using the protocol reported by Moses
and co-workers.23
1,3-Dipolar cycloaddition occurred when terminal
acetylene 26 reacted with aryl azides 28a-c using CuSO4.
H2O, sodium ascorbate and CH2Cl2/H2O 1:1 as the solvent,
yielding the ester-triazole compounds 29a-c in 80 to 89%
yield.24
Subsequent hydrolysis of 29a-c with NaOH in
EtOH/THF produced triazole-carboxylic acid retinoids 7-9
with 78 to 95% yield (Scheme 1).25
To synthesize the 1,2,3-triazole core, it was used a
click chemistry approach.18 Some important characteristics
of click reactions in organic synthesis are their modular
nature, high yields of products, broad scope, ability to
isolate products without chromatographic methods, use
of solvents with low toxicity, and stereospecificity, which
makes these methods very useful in organic synthesis.18
Another important characteristic of the click chemistry
approach in medicinal chemistry is the possibility to rapidly
synthesize a library of compounds with broad structural
diversification in order to obtain new molecules with
improved biological activity.19
The synthetic procedures for obtaining compounds 7-9
are shown in Scheme 1. The construction of 1,2,3-triazole
cores in click reactions depends on the reaction between
two building blocks, terminal acetylenes and azides.18,19
The synthesis was initiated by the reaction of
2,5-dimethyl-2,5-hexanediol 19 with 37% hydrochloric
acid for 64 h at room temperature, generating 2,5-dichloro-
2,5-dimethyl hexane 20 in 85% yield (Scheme 1).6
Then, Friedel-Crafts alkylation of 20 with an excess
of benzene 21 using AlCl3 as a catalyst provided
The 1,2,3-triazole retinoid derivatives 10-16 with
Scheme 1. Synthesis of 7-9. Reagents and conditions: (a) HCl 37%, rt, 64 h, 85%; (b) AlCl3, reflux, 48 h, 82%; (c) NBS, TsOH, CH3OH, reflux, 48 h,
79%; (d) 2-methyl-3-butyn-2-ol 24, PdCl2(PPh3)2, CuI, Et3N, reflux, 24 h, 81%; (e) KOH, toluene, reflux, 24 h, 70%; (f) t-BuONO, CH3CN, 15 min, 0 °C;
then TMSN3, rt, 5 h; (g) CuSO4.5H2O, CH2Cl2/H2O, sodium ascorbate, rt, 24 h; (h) NaOH, EtOH/THF, 18 h.

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Design, Synthesis and Anticancer Biological Evaluation of Novel 1,4-Diaryl-1,2,3-triazole
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Scheme 2. Synthesis of 10-16. Reagents and conditions: (a) 27d-i; then t-BuONO, CH3CN, 15 min, 0 °C; then TMSN3, rt, 5-24 h; (b) CuSO4.5H2O,
CH2Cl2/H2O, sodium ascorbate, rt, 48 h; (c) 15, then Fe0/CaCl2/EtOH, reflux, 48 h, 95%.
molecular modification in the aryl carboxylic unit were
synthesized by reaction of the terminal acetylene 26 and
aryl azides 28d-i with different aromatic substitution
patterns (Scheme 2).
The positional isomer 17 was also synthesized to
compare its biological activity with that of 7 (Scheme 3).
The synthesis of triazole 17 was initiated by nitration 21
of 1,1,4,4-tetramethyl-1,2,3,4-tetrahydronaphthalene 22,
generating nitro tetrahydronaphthalene 30,26 which
was reduced to the corresponding amine 31 using the
Fe0/CaCl2/EtOH system.27 Compound 31 was subsequently
transformed into the corresponding tetrahydronaphthalene
azide 32 in 78% yield using t-BuONO/TMSN3
(Scheme 3).23
Scheme 4. Synthesis of 35. Reagents and conditions: (a) 2-methyl-
3-butyn-2-ol 25, PdCl2(PPh3)2, CuI, Et3N, reflux, 24 h, 70%; (b) NaH,
hexane, reflux, 48 h, 65%.
T h e 1 , 3 - d i p o l a r cy c l o a d d i t i o n o f a r y l
tetrahydronaphthalene azide 32 and terminal acetylene 35
afforded the ester triazole compound 36 in 85% yield.24
Hydrolysis reaction of 36 with NaOH generated the
positional isomer 17 in 93% yield (Scheme 5).25
Finally, compound 18 was synthesized using Fokin’s
methodology29 by reaction between terminal acetylene 27
and azide ester 29a with 3 equiv. of potassium tert-butoxide
in dimethyl sulfoxide (DMSO) in order to compare the
anticancer activity of compound 18 with those of 7
and 17.
Scheme 3. Synthesis of 32. Reagents and conditions: (a) HNO3, Ac2O,
rt, 24 h, 92%; (b) Fe0, CaCl2, EtOH, reflux, 24 h, 75%; (c) t-BuONO,
CH3CN, 15 min, 0 °C, then TMSN3, rt, 12 h, 78%.
Unlike Fokin and co-workers,29 an excess of t-BuOK
was used, allowing one-pot preparation of compound 18
without requiring a deprotection step.
Next, the acetylene alcohol 34 was obtained in 90%
yield from the Sonogashira cross-coupling-type reaction
between the ethyl 4-bromobenzoate 33 and 2-methyl-
3-butyn-2-ol 24 using a PdCl2(PPh3)2/CuI system and
Et3N as base.20,22 The terminal acetylene 35 was obtained
in 65% yield via retro-Favorski reaction of acetylene
alcohol 34 with excess NaH in hexane under reflux for
48 h (Scheme 4).28
Biological activity
All triazole retinoid analogues synthesized were
evaluated for antiproliferative activity (Table 1) against
human leukemia cells (HL-60), chronic myeloid leukemia
cells (K562), human renal cell carcinoma (786), colorectal

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Scheme 5. Synthesis of 17. Reagents and conditions: (a) CuSO4.5H2O, CH2Cl2/H2O, sodium ascorbate, rt, 48 h, 85%; (b) NaOH, EtOH/THF, 18 h, 93%.
Scheme 6. Synthesis of 18. Reagents and conditions: (a) t-BuOK, DMSO, 96 h, 40 °C, 75%.
Table 1. Structures and antiproliferative activities of the 1,2,3-triazole retinoid analogs 7-18
IC50a / μM
entry
1
Compound
HL-60
15.6
K562
39.2
786
HT-29
MCF-7
41.6
PC-3
66.7
NIH/3T3
34.2
4b,c
64.2
54.1
2
3
7
8
16.4
70.0
66.6
67.4
21.6
65.2
6.7
18.1
50.7
21.9
66.6
24.1
61.0
66.6
4
9
51.7
61.7
15.9
10.2
> 200
> 200
155.9
5
6
7
10
11
12
> 200
47.0
> 200
> 200
> 200
> 200
> 200
> 200
> 200
> 200
> 200
> 200
> 200
> 200
> 200
> 200
> 200
> 200
> 200
66.0
> 200

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Design, Synthesis and Anticancer Biological Evaluation of Novel 1,4-Diaryl-1,2,3-triazole
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Table 1. Structures and antiproliferative activities of the 1,2,3-triazole retinoid analogs 7-18 (cont.)
IC50a / μM
HT-29
entry
8
Compound
HL-60
> 200
K562
> 200
786
MCF-7
> 200
PC-3
NIH/3T3
> 200
13
14
15
16
17
> 200
> 200
> 200
> 200
11.3
> 200
9
> 200
112.3
91.2
> 200
146.0
95.8
> 200
> 200
9.5
> 200
> 200
10.5
> 200
> 200
21.7
78.5
> 200
20.3
10
11
12
39.7
76.5
66.5
19.5
> 200
75.8
17.4
13
18
49.3
> 200
66.7
97.6
41.5
79.4
71.9
aIC50 is the concentration required for 50% inhibition of cell growth; btamibarotene (AM80) was used as a positive control in the MTT assay; cIC50 values
for tamibarotene (16.1 μM) are reported in reference 26.
adenocarcinoma cells (HT-29), breast adenocarcinoma cells
(MCF-7), and prostatic adenocarcinoma cells (PC-3).30
Thus, this study aims to determine whether retinoids
7-18, containing the 1,2,3-triazole ring, also have a good
anticancer activity profile against the solid tumor cells 786,
HT-29, MCF-7 and PC-3, since recent studies have shown
the potential of tamibarotene analogues against various
cancers other than APL.31
The assays performed herein show that tamibarotene
(AM80) 4 (Table 1, entry 1) has a better anticancer activity
profile against the HL60 leukemic cells (15.6 μM)31 than on
the K562 leukemic cells (39.2 μM), and moderate activity
on solid tumor cells 786 (64.2 μM), HT-29 (54.1 μM),
MCF-7 (41.6 μM) and PC-3 (66.7 μM).
1,2,3-Triazolic compound 7 containing a carboxylic acid
group at the para position exhibited good antiproliferative
activity against HL60 (16.4 μM), weak against K562
(66.6 μM), very good against HT-29 (6.7 μM), and good
against MCF-7 (18 μM) and PC3 (21.9 μM) (Table 1, entry 2).
However, triazole 8, with a carboxylic acid at the meta
position, showed lower anti-cancer activity compared to
triazole 7 (carboxylic acid in the para position), ranging
from 50.7 to 70 μM for all cell lines tested (Table 1, entry 3).
The chloro triazole 9 (Table 1, entry 4) had moderate
activity against HL60 and K562 (51.7 and 61.7 μM), good
activity against 786 and HT-29 (15.9 and 10.2 μM) and no
activity against MCF-7 and PC3 cells. Compounds with
chloro at the 3-position, such as UVI2007 6, are known
to interact with the RARβ receptor selectively, with
decreased binding affinity to the RARα receptor.10 The
chloro introduction in 7 selectively increased its anticancer
activity for the 786 and HT-29 cells.
Compound 10, containing an SO2NH2 group at the para
position, which can be considered a bioisosteric substitute
for carboxylic acid,32 had poor activity against all cancer
cell lines tested (Table 1, entry 5).
This aligns with previous results, which demonstrated
that classic bioisosteric replacement of a carboxylic acid
with a sulfonamide group does not work well for retinoid
compounds with anticancer activities against APL.33

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115
Interestingly, in another study with retinoid-containing
bioisosteric exchange groups, compounds having the
-SO2NH2 group had good activity against ovarian cancer.34
Compound 11 (Table 1, entry 6), which contains a
methoxy group at the para position, similarly to 7, had
moderate activity against HL 60 cells (47.0 μM) and
no activity against K562, 786 HT-29, MCF-7 and PC-3
cells. Compounds 12-14 (Table 1, entries 7-9), containing
dimethoxy, trimethoxy and methylenedioxy groups, had no
activity (> 200 μM) for all cell lines tested. The introduction
of the extra methoxy groups in the meta position can explain
these results.
Compound 15 (Table 1, entry 10), containing an
electron-withdrawing nitro group at the para position, had
weak activity against HL 60 cells (112.3 μM), even less
activity against K562 (146 μM), and no activity against
786, HT-29, MCF-7, and PC-3 cells.
Compound 16 (Table 1, entry 11), containing an NH2
group at the para position, had weak activity against
HL60 (91.2 μM) and K562 cells (95.8 μM); very good
activity against the solid tumor cells 786 (9.5 μM), HT-29
(11.3 μM), and MCF-7 (10.5 μM); and good activity against
PC-3 (21.7 μM) cells, which indicates that the NH2 group
is a good substituent for solid tumor cancer cells.
Triazole 17 (Table 1, entry 12), a positional isomer
of compound 7, showed moderate antiproliferative
activity against HL60 (39.6 μM), weak activity with
K562 (76.6 μM) and 786 (66.5 μM) cells, good activity
with HT-29 cells (19.5 μM), no activity against MCF-7
(> 200 μM) and weak activity for PC-3 (75.8 μM).
The 1,5-diaryl retinoid derivative 18 (Table 1, entry 13)
showed moderate to weak biological anti-cancer activity
against HL60 (49.3 μM), 786 (66.7 μM), HT-29 (97.6 μM),
MCF-7 (41.5 μM) and PC-3 (79.4 μM) cells and no activity
against K562 cells (> 200 μM).
Conclusions
In summary, 12 novel 1,2,3-triazole retinoid 7-18
derivatives of tamibarotene (AM80) 4 were synthesized
and evaluated for in vitro anti-cancer activity.
Among the compounds synthesized, triazole 7 stands
out, because it is equipotent to tamibarotene 4 in relation
to the anticancer activity against HL60 cells (Table 1,
entries 1-2). With respect to solid tumor cells, triazole 7 is
more active for all cancer cells tested when compared to
tamibarotene 4, with prominent anticancer activity for the
HT-29 colorectal adenocarcinoma cell line (6.7 μM) and
with good selectivity index (SI = 3.6). Compound 9 showed
the lowest cytotoxicity among compounds synthesized.
Compound 16, an aminoretinoid triazole containing
an NH2 group at the para position, had good anticancer
activity against all solid tumor cells tested. 16 is the most
potent anti-cancer compound for breast cancer (MCF-7
cells) of this work, and new studies are being conducted to
obtain new analogues of 16 aiming to obtain more potent
compounds and with less cytotoxicity.
Positional isomer 17 had lower potency than triazole 7,
indicating that obtaining a positional isomer of 1,4-diaryl-
1,2,3-triazole compounds is a critical factor in studies about
biological anti-cancer activities.
Experimental
General remarks
All solvents were distilled before use according to the
standard procedure. All reactions were performed under
an atmosphere of dry nitrogen and monitored by thin
layer chromatography (TLC) using prepared plates (silica
gel 60 F254 on aluminum). The chromatograms were
examined under both 254 and 360 nm UV light or with the
developing agent ethanolic vanillin and heat. Flash column
chromatography was performed on silica gel 60 (particle
size 200-400 meshASTM, purchased fromAldrich, USA)
and eluted with hexane or hexane/ethyl acetate in different
ratios. Melting points were determined using Fisatom 430D
equipment. Infrared (IR) spectra were recorded on a Nicolet
In relation to selectivity index (SI), tamibarotene showed
SI = 2.2 for HL60 cells (Table 2, entry 1). Compound 7
showed good SI = 3.6 for HT-29 cells (Table 2, entry 2) and
16 showed SI = 2.2, 1.8, and 1.9 for 786, HT-29 and MCF-7,
respectively (Table 2, entry 4). Compound 9 showed the
lowest cytotoxicity (Table 1, entry 3), with SI = 9.8 and
15.2 for 786 and HT-29 cells, respectively.
Table 2. Selectivity index (SI)a for most active retinoids analogues
Selectivity index
entry Compound
NIH/3T3 vs. HL60 NIH/3T3 vs. K562
NIH/3T3 vs. 786
NIH/3T3 vs. HT29 NIH/3T3 vs. MCF-7 NIH/3T3 vs. PC-3
1
2
3
4
4b
7
2.2
1.5
NCc
NCc
NCc
NCc
NCc
NCc
NCc
1.1
9.8
2.2
NCc
NCc
NCc
3.6
1.3
1.1
9
15.2
1.8
NCc
1.9
NCc
NCc
16
aSI, selectivity index: IC50 on normal cells / IC50 on cancer cells; bpositive control: tamibarotene for fibroblast cells (NIH/3T3); cNC: not calculated.

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Design, Synthesis and Anticancer Biological Evaluation of Novel 1,4-Diaryl-1,2,3-triazole
J. Braz. Chem. Soc.
iS5 spectrometer from Thermo Scientific. The 1H and 13C
nuclear magnetic resonance (NMR) spectra were recorded
in CDCl3 solutions using a Bruker 75 MHz or 300 MHz
spectrometer, as noted. Chemical shifts (d) are expressed as
parts per million (ppm) downfield from tetramethylsilane
as the internal standard. HR-ESI-MS (high resolution
electrospray ionization mass spectrometry) measurements
were carried out on a quadrupole time-of-flight instrument
(UltrOTOF-Q, Bruker Daltonics, Billerica, MA).
saturated NaHCO3 solution and extracted with ethyl acetate
(3 × 30 mL). The organic phase was dried over MgSO4 and
the solvent removed under vacuum. The product was purified
by distillation at low pressure (0.5 mm Hg) at temperatures
between 90 and 145 °C. The product 23 was obtained as
1
colorless oil in 79% yield. H NMR (300 MHz, CDCl3)37
d 1.25 (s, 6H, 2CH3), 1.26 (s, 6H, 2CH3), 1.66 (s, 4H, 2CH2),
7.16 (d, 1H, J 8.5 Hz, Ar-H), 7.23 (dd, 1H, J 8.5, 2.1 Hz,
13
Ar-H), 7.40 (d, 1H, J 2.1 Hz, Ar-H); C NMR (75 MHz,
CDCl3) d 31.72, 31.88, 34.06, 34.46, 34.83, 34.86, 119.40,
Procedure for the preparation of 2,5-dichloro-2,5-dimethyl
hexane (20)35
128.44, 128.65, 129.42, 143.82, 147.36.
General procedure for the preparation of acetylene alcohols
A solution of 2,5-dimethyl-2,5-hexanediol 19
(2.0 mmol) in 37% hydrochloric acid (42 mmol) was stirred
at room temperature for 65 h. The mixture was extracted
with ethyl acetate and the organic phase was washed with
5% NaHCO3 and dried over anhydrous MgSO4. The solvent
was removed under reduced pressure. The product was
purified by flash chromatography and recrystallization
using hexane as solvent, which furnished 2,5-dichloro-
2,5-dimethyl hexane 20 as white crystals in 85% yield.
25 and 34
To a solution of the bromines 23 or 33 (3.0 mmol) in
triethylamine (4.5 mL mmol-1), under nitrogen atmosphere,
PdCl2(PPh3)2 (0.075 mmol), CuI (0.15 mmol) and 2-methyl-
3-butyn-2-ol 24 (11.0 mmol) were added. The reaction
mixture was stirred under reflux at 75 °C for 20 h. The
excess triethylamine was removed by distillation, and the
reaction was extracted with ethyl acetate, dried over MgSO4
and the solvent removed under vacuum.
1
mp 68 °C; H NMR (300 MHz, CDCl3) d 1.58 (s, 12H,
3CH3), 1.93 (s, 4H, 2CH2); 13C NMR (75 MHz, CDCl3)
d 32.52, 41.14, 70.35.
2-Methyl-4-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-
naphthalen-2-yl)-but-3-yn-2-ol (25)22
Procedure for the preparation of 1,1,4,4-tetramethyl-
1,2,3,4-tetrahydronaphthalene (22)36
The product was purified by flash chromatography
(hexane/ethyl acetate 99:1) to give 25 as a yellow crystal in
81% yield. mp 95 °C; 1H NMR (300 MHz, CDCl3) d 1.23
(s, 6H, 2CH3), 1.25 (s, 6H, 2CH3), 1.60 (s, 6H, 2CH3), 1.65
(s, 4H, 2CH2), 1.98 (s, 1H, OH), 7.14 (dd, 1H, J 8.1, 1.6 Hz,
Ar-H), 7.21 (d, 1H, J 8.1 Hz, Ar-H), 7.34 (d, 1H, J 1.6 Hz,
Ar-H); 13C NMR (75 MHz, CDCl3) d 31.56, 31.67, 31.72,
34.15, 34.26, 34.85, 34.92, 65.66, 82.56, 92.54, 119.56,
126.54, 128.66, 129.89, 145.41.
To a solution of 2,5-dichloro-2,5-dimethyl hexane 20
(2.0 mmol) in benzene 21 (2 mL mmol-1),AlCl3 (0.2 mmol)
was added and the reaction mixture was stirred under reflux
in nitrogen atmosphere for 64 h. Excess benzene 21 was
distilled off and the resulting mixture was poured into
distilled water. The mixture was extracted with ethyl acetate
and the organic phase was dried over MgSO4. The solvent
was removed under vacuum. The residue was purified by
distillation at low pressure (0.5 mm Hg), and the product 22
was collected between 90 and 120 °C as a colorless oil in
Ethyl 4-(3-hydroxy-3-methylbut-1-yn-1-yl) benzoate (34)38
The product was purified by flash chromatography on
silica gel using hexane as eluent, to give 34 as a yellow oil
in 70% yield. 1H NMR (300 MHz, CDCl3) d 1.35 (t, 3H,
J 7.2 Hz, CH3), 1.60 (s, 6H, 2CH3), 2.41 (s, 1H, OH), 4.34
(q, 2H, J 7.2 Hz, CH2), 7.42 (d, 2H, J 9 Hz, Ar-H), 7.94 (d,
2H, J 9 Hz, Ar-H); 13C NMR (75 MHz, CDCl3) d 14.23,
31.31, 61.12, 65.52, 81.39, 96.70, 127.35, 129.33, 129.81,
131.46, 166.06.
1
82% yield. H NMR (300 MHz, CDCl3) d 1.30 (s, 12H,
3CH3), 1.71 (s, 4H, 2CH2), 7.15 (m, 2H, Ar-H), 7.33 (m,
2H, Ar-H); 13C NMR (75 MHz, CDCl3) d 31.88, 34.19,
35.10, 125.52, 126.45, 144.74.
Procedure for the preparation of 6-bromo-1,1,4,4-tetra-
methyl-1,2,3,4-tetrahydronaphthalene (23)
General procedure for the preparation of 6-ethynyl-
To a solution of tetrahydronaphthalene 22 (20.0 mmol)
in MeOH (2.5 mL mmol-1), under nitrogen atmosphere, NBS
(1.5 mmol) and TsOH (0.15 mmol) were added. The reaction
mixture was refluxed at 55 °C for 48 h, then poured into
1,1,4,4-tetramethyl-1,2,3,4-tetrahydronaphthalene 2638,39
To a solution of acetylene alcohol 25 (3.0 mmol) in
toluene (9.0 mL mmol-1) under nitrogen atmosphere, KOH

Vol. 29, No. 1, 2018
Aleixo et al.
117
(9.0 mmol) was added. The reaction mixture was refluxed
at 110 °C for 20 h and excess toluene was removed by
distillation. The residue was extracted with ethyl acetate
and the organic phase was dried over MgSO4. The solvent
was removed under vacuum. The product was purified by
flash chromatography on silica gel using hexane as eluent.
The product 25 was obtained as a yellow oil in 70% yield.
1H NMR (300 MHz, CDCl3) d 1.25 (sl, 12H, 4CH3), 1.66
(s, 4H, 2CH2), 2.99 (s, 1H, CH), 7.23 (m, 2H, Ar-H), 7.43
(sl, 1H, Ar-H); 13C NMR (75 MHz, CDCl3) d 31.6, 31.7,
34.1, 34.3, 34.8, 34.9, 75.9, 84.2, 119.0, 126.6, 129.1,
130.5, 145.0, 146.0.
in acetic anhydride (1 mL mmol-1), under nitrogen
atmosphere, at 0 °C, a solution of HNO3 (0.05 M) in 1 mL
of acetic anhydride was added dropwise. The mixture was
stirred at room temperature for 2 h and then extracted
with ether. The organic phase was washed with a saturated
solution of Na2CO3, and dried over anhydrous MgSO4.
The solvent was removed under vacuum and the product
was purified by flash chromatography on silica gel using
hexane as eluent. The product 30 was obtained as a yellow
1
oil in 92% yield. H NMR (300 MHz, CDCl3) d 1.29 (s,
6H, 2CH3), 1.31 (s, 6H, 2CH3), 1.71 (s, 4H, CH2), 7.42 (d,
1H, J 8.7 Hz, Ar-H), 7.92 (dd, 1H, J 8.7, 2.4 Hz, Ar-H),
8.16 (d, 1H, J 2.4 Hz, Ar-H); 13C NMR (75 MHz, CDCl3)
d 31.51, 31.63, 34.48, 34.60, 34.71, 120.42, 121.84, 127.67,
144.71, 146.71, 152.73.
Procedure for the preparation of ethyl 4-ethynylbenzoate
(35)40
To a solution of ethyl 4-(3-hydroxy-3-methylbut-1-yn-
1-yl) benzoate 34 (1.0 mmol) in hexane (12 mL mmol-1),
NaH (1.8 mmol) was added. The reaction mixture was
refluxed under nitrogen atmosphere for 48 h. The reaction
was extracted with ethyl acetate and the organic phase was
dried over MgSO4. The solvent was removed under vacuum.
The product was purified by flash chromatography on silica
gel using hexane as eluent. The product 35 was obtained
as a yellow oil in 65% yield. 1H NMR (300 MHz, CDCl3)
d 1.37 (t, 3H, J 6 Hz, CH3), 3.20 (s, 1H, CH), 4.35 (q, 2H,
J 6 Hz, CH2), 7.52 (d, 2H, J 9 Hz, Ar-H), 7.97 (d, 2H,
J 9 Hz, Ar-H); 13C NMR (75 MHz, CDCl3) d 14.27, 61.18,
79.93, 82.83, 126.59, 129.40, 130.48, 132.01, 165.91.
Procedure for the preparation of 5,5,8,8-tetramethyl-
5,6,7,8-tetrahydronaphthalen-2-amine (31)42
To a solution of nitro-1,2,3,4-tetrahydronaphthalenes 30
(5.0 mmol) in 95% ethanol (35 mL mmol-1), powdered iron
(150 mmol) and CaCl2 (50 mmol) were added. The mixture
was refluxed at 78 °C for 48 h. The reaction mixture was
extracted with ethyl acetate and the organic phase dried over
anhydrous MgSO4. The solvent was then removed under
vacuum. The product was purified by flash chromatography
using hexane/ethyl acetate 90:10 as eluent. The product 31
was obtained as a brown crystal in 75% yield. mp 71 °C;
1H NMR (300 MHz, CDCl3) d 1.21 (s, 6H, 2CH3), 1.23 (s,
6H, 2CH3), 1.62 (s, 4H, CH2), 3.48 (ls, 2H, NH2), 6.61 (d,
1H, J 2.7 Hz, Ar-H), 6.50 (dd, 1H, J 8.1, 2.7 Hz, Ar-H),
7.08 (d, 1H, J 8.1 Hz, Ar-H); 13C NMR (75 MHz, CDCl3)
d 31.80, 32.00, 33.53, 34.20, 35.23, 112.88, 113.62, 127.37,
135.38, 143.58, 145.82.
Procedure for the preparation of ethyl 4-amino-
3-chlorobenzoate (27c)41
To a solution of ethyl 4-aminobenzoate (2.0 mmol) in
acetonitrile (2 mL mmol-1), NCS (2.05 mmol) was added.
The mixture was refluxed for 5 h. Extraction was performed
with ethyl acetate and the organic layer was washed with 5%
NaOH, dried over anhydrous MgSO4, and the solvent was
removed under reduced pressure. The product was purified
by crystallization in hexane, which furnished 4-amino-
3-chlorobenzoate 27c as a purple crystal (mp 81-83 °C)
General procedure for the preparation of the azides 28a-i
and 3223
To a solution of amine 27a-i and 31 (2.0 mmol) in
acetonitrile (3.8 mL mmol-1), t-BuONO (4.3 mmol) and
TMSN3 (3.26 mmol) were added dropwise under a nitrogen
atmosphere, at 0 °C. The reaction mixture was stirred at
room temperature for 2 h. The solution was poured into
water, extracted with ethyl acetate and the organic phase
was dried over anhydrous MgSO4. The solvent was removed
under vacuum.
1
in 86% yield. H NMR (300 MHz, CDCl3) d 1.33 (t, 3H,
J 7.0 Hz, CH3), 4.29 (q, 2H, J 7.0 Hz, CH2), 6.70 (d, 1H,
J8.5 Hz,Ar-H), 7.72 (dd, 1H, J8.4, 1.9 Hz,Ar-H), 7.92 (d, 1H,
J 1.9 Hz,Ar-H); 13C NMR (75 MHz, CDCl3) d 14.30, 60.62,
114.35, 118.06, 120.63, 129.52, 131.12, 146.97, 165.74.
Procedure for the preparation of 1,1,4,4-tetramethyl-6-nitro-
Ethyl 4-azidobenzoate (28a)43
1,2,3,4-tetrahydronaphthalene (30)42
The compound was used without purification and it was
obtained as an orange oil in 96% yield. 1H NMR (300 MHz,
CDCl3) d 1.37 (t, 3H, J 7.1 Hz, CH3), 4.34 (q, 2H, J 7.0 Hz,
To a solution of tetrahydronaphthalene 22 (10 mmol)

118
Design, Synthesis and Anticancer Biological Evaluation of Novel 1,4-Diaryl-1,2,3-triazole
J. Braz. Chem. Soc.
2CH2), 7.04 (d, 2H, J 8.6 Hz, Ar-H), 8.02 (d, 2H, J 8.6 Hz,
Ar-H); 13C NMR (75 MHz, CDCl3) d 14.31, 61.04, 118.76,
127.01, 131.34, 144.59, 165.81.
(s, 6H, 2OCH3), 6.49 (d, 1H, J 2.5 Hz, CH2), 6.58 (dd, 1H,
J 8.4, 2.4 Hz,Ar-H), 6.81 (d, 1H, J 8.6 Hz,Ar-H); 13C NMR
(75 MHz, CDCl3) d 55.92, 56.17, 103.15, 110.40, 112.01,
132.72, 146.48, 149.95.
Ethyl 3-azidobenzoate (28b)44
The product was purified by flash chromatography on
silica gel using a hexane/ethyl acetate (94:6) solution as the
mobile phase, which furnished the product as a yellow oil
in 80% yield. 1H NMR (300 MHz, CDCl3) d 1.38 (t, 3H,
J 6 Hz, CH3), 4.36 (q, 2H, J 6 Hz, CH2), 7.18 (ddd, 1H,
J 8.0, 2.4, 1.0 Hz, Ar-H), 7.41 (dd, J 8.1, 7.7 Hz, Ar-H),
7.69 (dd, 1H, J 2.4, 1.6 Hz,Ar-H), 7.80 (ddd, 1H, J 7.6, 1.3,
1.1 Hz, Ar-H); 13C NMR (75 MHz, CDCl3) d 14.38, 61.35,
119.91, 123.28, 125.96, 129.76, 132.24, 140.49, 165.70.
5-Azido-1,2,3-trimethoxybenzene (28g)46
The product was purified by flash chromatography on
silica gel using a hexane/ethyl acetate (90:10) solution as
the mobile phase, which furnished the product as yellow
crystals in 87% yield. 1H NMR (300 MHz, CDCl3) d 3.78
(s, 3H, OCH3), 3.82 (s, 6H, 2OCH3), 6.21 (s, 2H, Ar-H);
13C NMR (75 MHz, CDCl3) d 56.17, 61.00, 96.40, 135.35,
135.64, 154.07.
5-Azidobenzo-1,3-dioxole (28h)46
Ethyl 4-azido-2-chlorobenzoate (28c)
The product was purified by flash chromatography
on silica gel using hexane as the mobile phase, which
furnished the product as a brown oil in 75% yield. 1H NMR
(300 MHz, CDCl3) d 5.95 (s, 2H, CH2), 6.47 (dd, 1H, J 8.2,
2.3 Hz, Ar-H), 6.51 (d, 1H, J 2.1 Hz, Ar-H), 6.75 (d, 1H,
J 8.3 Hz, Ar-H); 13C NMR (75 MHz, CDCl3) d 100.66,
101.59, 108.75, 111.58, 133.74, 145.01, 148.64.
The product was purified by flash chromatography
on silica gel using hexane as eluent, which furnished
the product as a yellow crystal in 92% yield. IR (KBr)
ν / cm-1 3257, 3102-2911, 2130, 1716, 1596, 1490, 1405,
1311-1286, 1247, 759; 1H NMR (300 MHz, CDCl3) d 1.37
(t, 3H, J 7.1 Hz, CH3), 4.35 (q, 2H, J 7.1 Hz, CH2), 7.20 (d,
1 H, J 8.4 Hz, Ar-H), 7.94 (dd, 1H, J 8.4, 1.8 Hz, Ar-H),
8.03 (d, 1H, J 1.8 Hz, Ar-H); 13C NMR (75 MHz, CDCl3)
d 14.28, 61.45, 119.23, 124.87, 127.92, 129.17, 132.00,
141.62, 164.78. HRMS (ESI+) m/z calcd. for C9H8ClN3O2
[M + H]+: 226.0383; found: 226.0394.
1-Azido-4-nitrobenzene (28i)47
The product was purified by flash chromatography on
silica gel using hexane as the mobile phase, which furnished
1
the product as yellow crystals in 79% yield. H NMR
(300 MHz, CDCl3) d 7.12 (d, 2H, J 9.0 Hz, Ar-H), 8.22 (d,
2H, J 9.0 Hz,Ar-H); 13C NMR (75 MHz, CDCl3) d 119.39,
125.61, 144.64, 146.87.
4-Azidobenzenesulfonamide (28d)45
The product was purified by flash chromatography on
silica gel using a hexane/ethyl acetate (60:40) solution as
eluent, which furnished the product as white crystals in
50% yield. 1H NMR (300 MHz, DMSO-d6) d 6.43 (sl, 2H,
NH2), 6.90 (dd, J 8.7, 0.5 Hz, Ar-H), 7.70 (dd, 2H, J 8.7,
0.5 Hz, Ar-H); 13C NMR (75 MHz, DMSO-d6) d 119.52,
127.66, 140.52, 142.98.
6-Azido-1,1,4,4-tetramethyl-1,2,3,4-tetrahydro-naphthalene
(32)
The product was purified by flash chromatography on
silica gel using hexane as the mobile phase, which furnished
the product as a brown oil in 86% yield. IR (KBr) ν / cm-1
2962-2861, 2111, 1604, 1517, 1496, 1309, 1276, 1139, 811;
1H NMR (300 MHz, CDCl3) d 1.24 (s, 6H, 2CH3), 1.25 (s,
6H, 2CH3), 1.66 (s, 4H, CH3), 6.81 (dd, 1H, J 8.4, 2.4 Hz,
Ar-H), 6.90 (d, 1H, J 2.4 Hz, Ar-H), 7.27 (d, 1H, J 8.4 Hz,
Ar-H); 13C NMR (75 MHz, CDCl3) d 31.74, 31.86, 34.51,
34.96, 35.28, 116.46, 116.90, 128.08, 137.01, 141.89,
146.83. HRMS (ESI+) m/z calcd. for C14H19N3 [M + H]+:
230.1657; found: 230.2411.
1-Azido-4-methoxybenzene (28e)46
The product was purified by flash chromatography on
silica gel using a hexane/ethyl acetate (95:5) solution as
the mobile phase, which furnished the product as yellow
crystals in 73% yield. 1H NMR (300 MHz, CDCl3) d 3.78
(s, 3H, OCH3), 6.87 (d, 2H, J 9.0 Hz, Ar-H), 6.94 (d, 2H,
J 8.9 Hz, Ar-H); 13C NMR (75 MHz, CDCl3) d 55.49,
115.05, 119.93, 132.26, 156.93.
General procedure for the preparation of triazole analogues
4-Azido-1,2-dimethoxybenzene (28f)46
10-15, 29a-c, 36
The product was purified by flash chromatography on
silica gel using a hexane/ethyl acetate (70:30) solution as
the mobile phase, which furnished the product as yellow
crystals in 78% yield. 1H NMR (300 MHz, CDCl3) d 3.84
To a solution of the azides (2.2 mmol) and terminal
acetylenes (2.0 mmol) in CH2Cl2/H2O (1:1; 4 mL mmol-1),
CuSO4.5H2O (0.15 mmol) and sodium ascorbate

Vol. 29, No. 1, 2018
Aleixo et al.
119
(0.35 mmol) were added. The mixture was stirred at room
temperature for 48 h and then extracted with ethyl acetate,
the organic phase was dried over anhydrous MgSO4 and
the solvent was removed under vacuum.
7.17 (d, 1H, J 8.6 Hz, Ar-H), 7.47 (m, 3H, Ar-H), 7.67 (dd,
1H, J 8.1, 1.4 Hz,Ar-H), 7.85 (d, 1H, J 1.4 Hz,Ar-H), 9.18
(s, 1H, *Tr-H); 13C NMR (75 MHz, DMSO-d6) d 31.51,
31.62, 33.88, 33.99, 34.50, 34.57, 55.83, 55.93, 104.53,
112.05, 119.16, 122.72, 123.26, 126.96, 127.61, 130.14,
144.43, 144.92, 147.35, 148.87, 149.34. HRMS (ESI+)
m/z calcd. for C24H29N3O2 [M + H]+: 392.2338; found:
392.2357. Calcd. for C24H29N3O2 [M + Na]+: 414.2158;
found: 414.2173. *Tr: triazole hydrogen.
4-(4-(5,5,8,8-Tetramethyl-5,6,7,8-tetrahydro-naphtha-lene-
2-yl)-1H-1,2,3-triazol-1-yl)benzene-sulfonamide (10)
The product 10 was purified by crystallization in an
ethyl acetate/ethanol (70:30) solution and then filtered
using Celite. The product was obtained as white crystals
in 50% yield. mp 282 °C; IR (KBr) ν / cm-1 3359, 3162,
3066, 2964-2859, 1598, 1482, 1348, 1159, 844; 1H NMR
(300 MHz, DMSO-d6) d 1.27 (s, 6H, 2CH3), 1.32 (s, 6H,
2CH3), 1.67 (s, 4H, CH2), 7.44 (d, 1H, J 8.3 Hz, Ar-H),
7.55 (s, 2H, Ar-H), 7.69 (dd, 1H, J 8.2, 1.6 Hz, Ar-H), 7.87
(d, 1H, J 1.6 Hz, Ar-H), 8.06 (d, 2H, J 8.8 Hz, Ar-H), 8.18
(d, 2H, J 8.8 Hz, Ar-H), 9.40 (s, 1H, *Tr-H); 13C NMR
(75 MHz, DMSO-d6) d 31.58, 31.69, 34.01, 34.10, 34.57,
119.41, 120.15, 122.85, 123.57, 127.20, 127.64, 138.73,
143.81, 144.91, 145.16, 148.03. HRMS (ESI+) m/z calcd.
for C22H26N4O2S [M + H]+: 411.1854; found: 411.1862.
Calcd. for C22H26N4O2S [M + Na]+: 433.1674; found:
433.1679. *Tr: triazole hydrogen.
4-(5,5,8,8-Tetramethyl-5,6,7,8-tetrahydronaphthalene-2-yl)-
1-(3,4,5-trimethoxyphenyl)-1H-1,2,3-triazole (13)
The product 13 was purified by flash chromatography on
silica gel using a hexane/ethyl acetate (85:15) solution as the
mobile phase, which furnished the product as white crystals
in 84% yield. mp 99 °C; IR (KBr) ν / cm-1 2959-2857, 1604,
1518, 1261, 1137, 1043, 1027, 797; 1H NMR (300 MHz,
DMSO-d6) d 1.25 (s, 6H, 2CH3), 1.30 (s, 6H, 2CH3), 1.65 (s,
4H, 2CH2), 3.71 (s, 3H, OCH3), 3.88 (s, 6H, 2OCH3), 7.23
(s, 2H, Ar-H), 7.42 (d, 1H, J 8.3 Hz, Ar-H), 7.65 (dd, 1H,
J 8.3, 1.7 Hz, Ar-H), 7.83 (d, 1H, J 1.7 Hz, Ar-H), 9.18 (s,
1H, *Tr-H); 13C NMR (75 MHz, DMSO-d6) d 31.68, 31.81,
34.08, 34.18, 34.65, 34.72, 56.52, 60.44, 98.13, 119.49,
122.99, 123.44, 127.25, 127.59, 132.73, 137.55, 144.83,
145.22, 147.66, 153.73. HRMS (ESI+) m/z calcd. for
C25H31N3O3 [M + H]+: 422.2443; found: 422.2448. Calcd.
for C25H31N3O3 [M + Na]+: 444.2263; found: 444.2267.
*Tr: triazole hydrogen.
1-(4-Methoxyphenyl)-4-(5,5,8,8-tetramethyl-5,6,7,8-tetra-
hydronaphthalene-2-yl)-1H-1,2,3-triazole (11)
The product 11 was purified by flash chromatography on
silica gel using a hexane/ethyl acetate (80:20) solution as the
mobile phase, which furnished the product as white crystals
in 81% yield. mp 135 °C; IR (KBr) ν / cm-1 2960-2856, 1518,
1253, 1042, 832, 781; 1H NMR (300 MHz, DMSO-d6) d1.26
(s, 6H, 2CH3), 1.30 (s, 6H, 2CH3), 1.66 (s, 4H, 2CH2), 3.83
(s, 3H, OCH3), 7.16 (d, 2H, J 9.0 Hz, Ar-H), 7.41 (d, 1H,
J 8.2 Hz, Ar-H), 7.65 (d, 1H, J 8.2 Hz, Ar-H), 7.85 (m, 3H,
Ar-H), 9.17 (s, 1H, *Tr-H); 13C NMR (75 MHz, DMSO-d6)
d 31.56, 31.66, 33.94, 34.05, 34.62, 55.63, 114.95, 119.20,
121.60, 122.75, 123.39, 127.04, 127.63, 130.16, 144.51,
145.00, 147.48, 159.29. HRMS (ESI+) m/z calcd. for
C23H27N3O [M + H]+: 362.2232; found: 262.2246. Calcd.
for C23H27N3O [M + Na]+: 384.2052; found: 384.2066. *Tr:
triazole hydrogen.
1-(Benzo-1,3-dioxol-5-yl)-4-(5,5,8,8-tetramethyl-
5,6,7,8-tetrahydronaphtha-lene-2-yl)-1H-1,2,3-triazole (14)
The product 14 was purified by crystallization in a
hexane/ethyl acetate (70:30) solution. The product was
obtained as white crystals in 69% yield. mp 163 °C; IR
(KBr) ν / cm-1 3140, 2957-2857, 1505, 1485, 1248, 1040,
804; 1H NMR (300 MHz, DMSO-d6) d 1.27 (s, 6H, 2CH3),
1.31 (s, 6H, CH3), 1.67 (s, 4H, 2CH2), 6.16 (s, 2H, CH2), 7.13
(d, 1H, J 8.3 Hz, Ar-H), 7.42 (d, 1H, J 8.3 Hz, Ar-H), 7.43
(dd, 1H, J 8.3, 1.9 Hz, Ar-H), 7.53 (d, 1H, J 2.1 Hz, Ar-H),
7.65 (dd, 1H, J 8.2, 1.7 Hz, Ar-H), 7.83 (d, 1H, J 1.6 Hz,
Ar-H), 9.13 (s, 1H, *Tr-H); 13C NMR (75 MHz, DMSO-d6)
d 31.55, 31.66, 33.94, 34.05, 34.55, 34.61, 101.84, 102.21,
108.73, 113.69, 119.41, 122.73, 123.41, 127.06, 127.53,
131.18, 144.58, 145.02, 147.44, 148.27. HRMS (ESI+)
m/z calcd. for C23H25N3O2 [M + H]+: 376.2025; found:
376.2032. Calcd. for C23H25N3O2 [M + Na]+: 398.1845;
found: 398.1846. *Tr: triazole hydrogen.
1-(3,4-Dimethoxyphenyl)-4-(5,5,8,8-tetramethyl-
5,6,7,8-tetrahydronaphtha-lene-2-yl)-1H-1,2,3-triazole (12)
The product 12 was purified by flash chromatography
on silica gel using a hexane/ethyl acetate (85:15) solution
as eluent, which furnished the product as white crystals in
62% yield. mp 126 °C; IR (KBr) ν / cm-1 2958-2857, 1603,
1510, 1470, 1232, 1125, 1040, 817; 1H NMR (300 MHz,
DMSO-d6) d 1.27 (s, 6H, 2CH3), 1.32 (s, 6H, 2CH3), 1.68
(s, 4H, 2CH2), 3.84 (s, 3H, OCH3), 3.89 (s, 3H, OCH3),
1-(4-Nitrophenyl)-4-(5,5,8,8-tetramethyl-5,6,7,8-tetra-
hydronaphthalen-2-yl)-1H-1,2,3-triazole (15)
The product 15 was purified by flash chromatography

120
Design, Synthesis and Anticancer Biological Evaluation of Novel 1,4-Diaryl-1,2,3-triazole
J. Braz. Chem. Soc.
on silica gel using a hexane/ethyl acetate (90:10) solution
as the mobile phase, which furnished the product as yellow
crystals in 61% yield. mp 205 °C; IR (KBr) ν / cm-1 3157,
3096, 2965-2861, 1596, 1523, 1338, 1231, 1037, 849, 748;
1H NMR (300 MHz, CDCl3) d 1.31 (s, 6H, 2CH3), 1.35 (s,
6H, 2CH3), 1.71 (s, 4H, 2CH2), 7.40 (d, 1H, J 8.3 Hz,Ar-H),
7.61 (dd, 1H, J 8.2, 1.7 Hz, Ar-H), 7.87 (d, 1H, J 1.7 Hz,
Ar-H), 8.03 (d, 2H, J 9.0 Hz,Ar-H), 8.24 (s, 1H, *Tr-H), 8.42
(d, 2H, J 9.0 Hz,Ar-H); 13C NMR (75 MHz, CDCl3) d 31.79,
31.86, 34.35, 34.43, 34.94, 35.05, 116.72, 120.31, 123.31,
124.18, 125.54, 126.65, 127.32, 141.26, 145.79, 146.05,
147.10, 149.60. HRMS (ESI+) m/z calcd. for C22H24N4O2
[M + H]+: 377.1977; found: 377.1974. *Tr: triazole hydrogen.
2H, J 8.8 Hz, Ar-H), 8.21 (d, 2H, J 8.8 Hz, Ar-H), 8.21 (s,
1H, *Tr-H); 13C NMR (75 MHz, CDCl3) d 14.31, 31.80,
31.86, 34.32, 34.42, 34.98, 35.10, 61.43, 116.86, 119.77,
123.29, 124.12, 127.08, 127.23, 130.46, 131.30, 140.12,
145.67, 145.69, 149.11, 165.48. HRMS (ESI+) m/z calcd.
for C25H29N3O2 [M + H]+: 404.2338; found: 404.2346. *Tr:
triazole hydrogen.
Ethyl 3-(4-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-
naphthalen-2-yl)-1H-1,2,3-triazol-1-yl)benzoate (29b)
The product 29b was purified by crystallization from
hexane and was obtained as white crystals in 89% yield.
mp 146 °C; IR (KBr) ν / cm-1 3145, 2989-2863, 1708,
1590, 1481, 1363, 1278, 1241, 1186, 1122, 1035, 892, 759;
1H NMR (300 MHz, CDCl3) d 1.30 (s, 6H, 2CH3), 1.35 (s,
6H, 2CH3), 1.42 (t, 3H, J 7.0 Hz, CH2), 1.71 (s, 4H, 2CH2),
4.43 (q, 2H, J 7.0 Hz, CH2), 7.39 (d, 1H, J 8.2 Hz, Ar-H),
7.63 (m, 2H, Ar-H), 7.87 (d, 1H, J 1.8 Hz, Ar-H), 8.10 (m,
2H,Ar-H), 8.22 (s, 1H, *Tr-H), 8.38 (dd, 1H, J 8.1, 1.7 Hz,
Ar-H); 13C NMR (75 MHz, CDCl3) d 14.32, 31.81, 31.86,
34.32, 34.42, 34.99, 35.10, 61.66, 117.15, 120.99, 123.27,
124.09, 124.69, 127.22, 129.49, 129.98, 132.22, 137.27,
145.59, 145.64, 149.01, 165.42. HRMS (ESI+) m/z calcd.
for C25H29N3O2 [M + H]+: 404.2338; found: 404.2348. *Tr:
triazole hydrogen.
Procedure for the preparation of 4-(4-(5,5,8,8-tetramethyl-
5,6,7,8-tetrahydronaphthalen2-yl)-1H-1,2,3-triazol-1-yl)
aniline (16)
Toasolutionof1-(4-nitrophenyl)-4-(5,5,8,8-tetramethyl-
5,6,7,8-tetrahydronaphthalen-2-yl)-1H-1,2,3-triazole 15
(1 mmol) in 95% ethanol (35 mL mmol-1), powdered
iron (30 mmol) and CaCl2 (10 mmol) were added. The
mixture was stirred under reflux for 48 h. Extraction was
performed with ethyl acetate and the organic phase was
dried over anhydrous MgSO4. The solvent was removed
under vacuum. There was no need for purification, and the
product was obtained as yellow crystals in 95% yield. mp
88 °C; IR (KBr) ν / cm-1 3157, 3096, 2965-2861, 1596,
Ethyl 3-chloro-4-(4-(5,5,8,8-tetramethyl-5,6,7,8-tetra-
hydronaphthalen-2-yl)-1H-1,2,3-triazol-1-yl)benzoate (29c)
The product 29c was purified by crystallization from
hexane and was obtained as white crystals in 80% yield.
mp 121 °C; IR (KBr) ν / cm-1 3143, 3056, 2969-2863,
1716, 1616, 1502, 1446, 1363, 1268, 1105, 1039, 771;
1H NMR (300 MHz, CDCl3) d 1.30 (s, 6H, 2CH2), 1.34 (s,
6H, 2CH2), 1.42 (t, 3H, J 7.1 Hz, CH3), 1.71 (s, 4H, 2CH2),
4.42 (q, 2H, J 7.2 Hz, CH2), 7.38 (d, 1H, J 8.1 Hz, Ar-H),
7.59 (dd, 1H, J 8.1, 1.8 Hz, Ar-H), 7.80 (d, 1H, J 8.3 Hz,
Ar-H), 7.89 (d, 1H, J 1.7 Hz, Ar-H), 8.10 (dd, 1H, J 8.3,
1.8 Hz, Ar-H), 8.24 (s, 1H, *Tr-H), 8.25 (d, 1H, J 1.6 Hz,
Ar-H); 13C NMR (75 MHz, CDCl3) d 14.26, 31.81, 31.85,
34.31, 34.42, 34.98, 35.10, 61.95, 120.94, 123.33, 124.12,
127.07, 127.22, 127.50, 128.12, 129.03, 132.07, 132.55,
138.14, 145.62, 145.68, 148.22, 164.36. HRMS (ESI+)
m/z calcd. for C25H28ClN3O2 [M + H]+: 438.1948; found:
438.1939. *Tr: triazole hydrogen.
1
1523, 1338, 1231, 1037, 849, 748; H NMR (300 MHz,
CDCl3) d 1.29 (s, 6H, 2CH3), 1.33 (s, 6H), 1.70 (s, 4H),
6.79 (d, 2H, J 8.8 Hz, Ar-H), 7.36 (d, 1H, J 8.1 Hz, Ar-H),
7.52 (d, 2H, J 8.7 Hz, Ar-H), 7.57 (dd, 1H, J 8.1, 1.8 Hz,
Ar-H), 7.85 (d, 1H, J 1.8 Hz, Ar-H), 8.01 (s, 1H, *Tr-H);
13C NMR (75 MHz, CDCl3) d 31.82, 31.85, 34.27, 34.40,
35.01, 35.13, 115.30, 117.47, 122.27, 123.20, 123.97,
127.11, 127.66, 128.80, 145.19, 145.51, 147.00, 148.30.
HRMS (ESI+) m/z calcd. for C22H26N4: 347.2235; found:
347.2130. *Tr: triazole hydrogen.
Ethyl 4-(4-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-
naphthalen-2-yl)-1H-1,2,3-triazol-1-yl)benzoate (29a)
The product 29a was purified by flash chromatography
on silica gel using a hexane/ethyl acetate (98:2) solution as
the mobile phase, which furnished the product as yellow
crystals in 82% yield. mp 155 °C; IR (KBr) ν / cm-1 3149,
3070, 2962-2863, 1720, 1608, 1517, 1484, 1415, 1392,
1274, 1222, 1106, 1029, 844, 769, 690; 1H NMR (300 MHz,
CDCl3) d 1.30 (s, 6H, 2CH3), 1.34 (s, 6H, 2CH3), 1.41
(t, 3H, J 7.2 Hz, CH3), 1.71 (s, 4H, 2CH2), 4.41 (q, 2H,
J 7.2 Hz, CH2), 7.38 (d, 1H, J 8.2 Hz, Ar-H), 7.60 (dd, 1H,
J 8.2, 1.8 Hz, Ar-H), 7.87 (d, 1H, J 1.8 Hz, Ar-H), 7.89 (d,
Ethyl 4-(1-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-
naphthalen-2-yl)-1H-1,2,3-triazol-4-yl)benzoate (36)
The product was purified by crystallization in hexane,
providing 36 as yellow crystals in 85% yield. mp 155 °C;
IR (KBr) ν / cm-1 3143, 3056, 2969-2863, 1716, 1616, 1502,
1446, 1363, 1268, 1105, 1039, 771; 1H NMR (300 MHz,

Vol. 29, No. 1, 2018
Aleixo et al.
121
CDCl3) d 1.31 (s, 6H, 2CH3), 1.34 (s, 6H, 2CH3), 1.40
(t, 3H, J 6.0 Hz, CH3), 1.73 (s, 4H, 2CH2), 4.39 (q, 2H,
J 6.0 Hz, CH2), 7.46 (m, 2H), 7.68 (s, 1H), 7.98 (d, 2H,
J 8.4 Hz, Ar-H), 8.13 (d, 2H, J 8.4 Hz, Ar-H), 8.20 (s, 1H,
*Tr-H); 13C NMR (75 MHz, CDCl3) d 14.36, 31.80, 34.37,
34.69, 34.76, 34.79, 61.07, 118.07, 118.92, 125.54, 128.11,
130.07, 130.23, 134.64, 146.25, 147.05, 166.32. HRMS
(ESI+) m/z calcd. for C25H29N3O2 [M + H]+: 404.2338;
found: 404.2340. *Tr: triazole hydrogen.
127.27, 130.19, 131.29, 137.32, 145.69, 147.73, 149.07,
170.00. HRMS (ESI+) m/z calcd. for C23H25N3O2 [M + H]+:
376.2025; found: 376.2040. *Tr: triazole hydrogen.
3-Chloro-4-(4-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-
naphthalen-2-yl)-1H-1,2,3-triazol-1-yl) benzoic acid (9)
The product 9 was obtained as white crystals in 91%
yield. mp 241 °C; IR (KBr) ν / cm-1 3434, 2957-2858, 1691,
1604, 1436-1419, 1288, 1247, 1016; 1H NMR (300 MHz,
CDCl3) d 1.30 (s, 6H, 2CH3), 1.35 (s, 6H, 2CH3), 1.71
(s, 4H, 2CH2), 7.40 (d, 1H, J 8.3 Hz, Ar-H), 7.60 (dd,
1H, J 8.2, 1.9 Hz, Ar-H), 7.86 (d, 1H, J 8.2 Hz, Ar-H),
7.90 (d, 1H, J 1.7 Hz, Ar-H), 8.18 (dd, 1H, J 8.4, 1.9 Hz,
Ar-H), 8.27 (s, 1H, *Tr-H), 8.33 (d, 1H, J 1.8 Hz, Ar-H);
13C NMR (75 MHz, CDCl3) d 31.81, 31.85, 34.33, 34.44,
34.97, 35.09, 120.99, 123.38, 124.20, 126.85, 127.27,
127.69, 128.25, 129.69, 131.46, 132.75, 138.79, 145.75,
145.78, 148.30, 168.75. HRMS (ESI+) m/z calcd. for
C23H24ClN3O2 [M + H]+: 410.1635; found: 410.1630. *Tr:
triazole hydrogen.
General procedure for the preparation of acid triazole
analogues 7-9, 17
To a solution of 29a-c or 36 (1 mmol) in ethanol 96%
(7 mL mmol-1), NaOH (18.75 mmol) was added. The
mixture was stirred at room temperature for 24 h. To the
final solution, 37% HCl was added until pH = 2. Then the
solution was extracted with ethyl acetate. The organic phase
was dried over anhydrous MgSO4, and the solvent was then
removed under vacuum. The products were purified by
crystallization from hexane/ethyl acetate (70:30).
4-(1-(5,5,8,8-Tetramethyl-5,6,7,8-tetrahydro-naphtha-len-
2-yl)-1H-1,2,3-triazol-4-yl)benzoic acid (17)
4-(4-(5,5,8,8-Tetramethyl-5,6,7,8-tetrahydro-naphtha-len-
2-yl)-1H-1,2,3-triazol-1-yl)benzoic acid (7)
The product 17 was obtained as white crystals in 93%
yield. mp 219 °C; IR (KBr) ν /cm-1 3091-2534, 2961,
1694-1614, 1504, 1418, 1313, 1278, 1245, 1181, 1043,
1014, 965, 864, 844, 779, 713, 697; 1H NMR (300 MHz,
DMSO-d6) d 1.29 (s, 6H, 2CH3), 1.33 (s, 6H, 2CH3), 1.69 (s,
4H, 2CH2), 7.58 (d, 1H, J 8.4 Hz,Ar-H), 7.70 (dd, 1H, J 8.7,
2.4 Hz, Ar-H), 7.84 (d, 1H, J 2.4 Hz, Ar-H), 8.07 (s, 4H,
Ar-H), 9.42 (s, 1H, *Tr-H); 13C NMR (75 MHz, DMSO-d6)
d 31.47, 34.05, 34.42, 117.63, 117.92, 120.75, 125.30,
128.20, 130.13, 134.36, 134.51, 145.42, 146.26, 146.49,
167.06. HRMS (ESI+) m/z calcd. for C23H25N3O2 [M + H]+:
376.2025; found: 376.2019. *Tr: triazole hydrogen.
The product 7 was obtained as white crystals in 95%
yield. mp 278 °C; IR (KBr) ν / cm-1 3900-3500, 2961-2924,
1699-1608, 1558-1410, 1275, 1037, 978, 860, 806, 772;
1H NMR (300 MHz, DMSO-d6) d1.27 (s, 6H, 2CH3), 1.32 (s,
6H, 2CH3), 1.67 (s, 4H, 2CH2), 7.44 (d, 1H, J 8.4 Hz,Ar-H),
7.70 (dd, 1H, J 8.4, 1.4 Hz, Ar-H), 7.89 (d, 1H, J 1.4 Hz,
Ar-H), 8.11 (d, 2H, J 8.7 Hz, Ar-H), 8.18 (d, 2H, J 8.7 Hz,
Ar-H), 9.43 (s, 1H, *Tr-H), 13.33 (s, 1H, OH); 13C NMR
(75 MHz, DMSO-d6) d 31.96, 32.07, 34.38, 34.48, 34.97,
35.02, 119.68, 120.03, 123.22, 123.94, 127.54, 127.67,
131.59, 140.02, 145.24, 145.51, 148.38, 167.03. HRMS
(ESI+) m/z calcd. for C23H25N3O2 [M + H]+: 376.2025; found:
376.2029. Calcd. for C23H25N3O2 [M + Na]+: 398.1845;
found: 398.1849. *Tr: triazole hydrogen.
General procedure for the preparation of acid triazole
analogue 18
3-(4-(5,5,8,8-Tetramethyl-5,6,7,8-tetrahydronaphtha-len-
2-yl)-1H-1,2,3-triazol-1-yl)benzoic acid (8)
To a solution of terminal acetylene 27 (1 mmol) and
aryl azide 29a (1 mmol) in DMSO (3 mL), potassium
t-butoxide (3 mmol) was added under nitrogen atmosphere.
The mixture was stirred for 96 h at 40 °C. Then, a 37%
HCl solution was added until pH = 3. The reaction mixture
was extracted with ethyl acetate and the organic phase was
dried over anhydrous MgSO4. The solvent was removed
under reduced pressure.
The product 8 was obtained as white crystals in 78%
yield. mp 187 °C; IR (KBr) ν / cm-1 3426, 2958-2859,
2667, 2572, 1737, 1710, 1691, 1594, 1465, 1363, 1311,
1245, 1230, 1105, 1054, 1049, 900, 829, 754, 705; 1H NMR
(300 MHz, CDCl3) d 1.30 (s, 6H, 2CH3), 1.35 (s, 6H,
2CH3), 1.71 (s, 4H, 2CH2), 7.40 (d, 1H, J 8.2 Hz, Ar-H),
7.66 (m, 2H, Ar-H), 7.89 (d, 1H, J 1.8 Hz, Ar-H), 8.19 (m,
2H, Ar-H), 8.26 (s, 1H), 8.46 (s, 1H, *Tr-H); 13C NMR
(75 MHz, CDCl3) d 31.81, 31.85, 34.32, 34.43, 34.99,
35.10, 117.28, 121.46, 123.34, 124.17, 125.52, 126.94,
4-(5-(5,5,8,8-Tetramethyl-5,6,7,8-tetrahydronaphtha-len-
2-yl)-1H-1,2,3-triazol-1-yl)ben-zoic acid (18)
The product 18 was obtained as yellow crystals in 75%

122
Design, Synthesis and Anticancer Biological Evaluation of Novel 1,4-Diaryl-1,2,3-triazole
J. Braz. Chem. Soc.
yield. IR (KBr) ν / cm-1 2956-2859, 1691, 1606, 1430, 1319,
1292, 1236, 1133, 1008, 970, 863, 825, 774; H NMR
concentrations of each compound: 0.25, 2.5, 25 and
250 μg mL-1 for 48 h. The adherent cells were then fixed
with 100 μL of 40% trichloroacetic acid (TCA), incubated
for 30 min at 4 °C and colored using sulforhodamine B
(SRB) according to the methodology described by Skehan
and co-workers.30 For non-adherent cells, at the end of the
exposure time to the substances, the medium was removed
and replaced with 0.2 mL of fresh MTT solution (final
concentration 0.5 mg mL-1). The plates were incubated
at 37 °C for 4 h. After the incubation, the medium was
removed and insoluble MTT crystals and formazan were
dissolved by adding 0.2 mL of DMSO and the optical
densities were read in a spectrophotometric plate reader
at 540 nm.
1
(300 MHz, DMSO-d6) d 0.95 (6H, s, 2CH3), 1.20 (s, 6H,
2CH3), 1.56 (s, 4H, 2CH2), 7.01 (d, 1H, J 1.5Hz, Ar-H),
7.17 (dd, 1H, J 8.1, 1.8 Hz, Ar-H), 7.38 (d, 1H, J 8.4 Hz,
Ar-H), 7.55 (d, 2H, J 8.4Hz, Ar-H), 8.08 (d, 2H, J 8.4Hz,
Ar-H), 8.15 (s, 1H, *Tr-H); 13C NMR (75 MHz, DMSO-d6)
d 31.53, 31.80, 34.19, 34.39, 34.64, 34.72, 123.43, 125.96,
126.40, 126.97, 127.67, 131.05, 132.44, 133.40, 138.43,
140.23, 145.27, 146.23, 166.92. HRMS (ESI+) m/z calcd.
for C23H25N3O2 [M + H]+: 376.2025; found: 376.2023. *Tr:
triazole hydrogen.
Cytotoxicity assay30,48,49
The concentrations of tamibarotene (Sigma-Aldrich®)
used as positive controls were 0.25, 2.5, 25 and 250 μg mL-1.
For all cells, three absorbance readings were obtained
at a wavelength of 540 nm at time zero (T0) and 48 h for
both the negative controls (C) and the cells treated with
the compounds (T). When C > T ≥ T0, growth (%) was
determined using the equation 100 × [(T − T0) / C – T0]
(cytostatic effect). When T < T0, growth (%) was
determined using the equation 100 × [(T − T0) / (T0)]
(cytocide effect) using Excel 2003/2007. The dose that
inhibited growth by 50% (IC50) was determined by non-
linear regression analyses in Origin 6.0 software.
Cell lines
Sixstrainsofhumantumorcellswereused:786-0(ATCC-
CRL-1932, kidney carcinoma), HT-29 (ATCC-HTB-38,
colon carcinoma), MCF-7 (ATCC-HTB-22, breast
adenocarcinoma), PC-3 (PC-3 ATCC-CRL-1435,
prostatic adenocarcinoma), HL-60 (ATCC-CCL-240,
promyelocytic leukemia) and K-562 (ATCC-CCL-243,
chronic myelogenous leukemia). The strains were donated
by Professor Dr João Ernesto de Carvalho (CPQBA-
UNICAMP) and stored under liquid nitrogen. For
cytotoxicity assays, the cells were thawed in RPMI-1640
medium supplemented with 10% fetal calf serum and 1%
streptomycin-penicillin, then maintained in an incubator
with 5% CO2 at 37 °C in a humid environment for growth.
Supplementary Information
Supplementary data are available free of charge at
http://jbcs.sbq.org.br as PDF file.
Sample preparation for analysis
The samples were prepared by adding DMSO
(0.1 g mL-1) and then diluted in complete medium to achieve
a maximum DMSO concentration of 0.25%.
Acknowledgments
This study was supported by grants from FUNDECT-MS
(Process number 23/200.012/2008 and 23/200.071/2010),
PROPP-UFMS, CNPq and CAPES. We thank Dr Janet
W. Reid (JWR Associates) for her assistance with English
corrections. Special thanks to the Laboratory of Natural
Products and Mass Spectrometry (LAPNEM) of the Federal
UniversityofMatoGrossodoSulfortheHPLC-DAD-MS/MS
analysis.
Cell preparation
For cell counting, the complete medium was aspirated
then 0.5 mL of EDTA-trypsin (1 mM EDTA, 0.25%
trypsin) in PBS, pH 7.4, was added. The cells were kept
in an incubator for 3-5 min. After centrifugation for 4 min
at 1000 rpm, trypsin and the medium were discarded
and fresh medium was added to obtain a cell suspension.
For non-adherent cells the suspension was obtained by
centrifugation.
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Submitted: April 6, 2017
Published online: June 27, 2017